Constitutive plasma membrane interaction of active Rho GEF Ect2 inhibits cortex contraction pulses

During cell division, multiple guanine nucleotide exchange factors (GEFs), including the mitotic regulator Ect2 and Lbc-type GEFs, regulate the dynamics of RhoA activity and actomyosin contractility. In interphase, Lbc-type GEFs are predominantly cytosolic and are transiently recruited to the plasma membrane via a Rho-dependent positive-feedback mechanism that generates stochastic pulses of Rho activity and cortical contractions. In contrast, during interphase, Ect2 is primarily sequestered in the nucleus away from the cytoplasm and the plasma membrane. Following nuclear envelope breakdown in mitosis, Ect2 is released into the cytosol and constitutively associates with the plasma membrane independently of Rho via a C-terminal polybasic cluster. Here, we mimicked this mitotic state by expressing a constitutively active Ect2 variant that is lacking its nuclear localization signal in interphase, to examine its interplay with Lbc-stimulated pulsatile Rho dynamics. We found that active cytosolic Ect2 suppressed pulsatile dynamics and promoted peripheral enrichment of Rho activity and myosin. Pulse suppression required Ect2 catalytic activity, Rho-dependent positive feedback, and constitutive plasma membrane association. In contrast, an Ect2 variant that was lacking constitutive plasma membrane association via the polybasic cluster, was not only incapable of pulse suppression but instead even stimulated strong pulsatile Rho dynamics. Together, our results identify constitutive plasma membrane association as a key distinction between Lbc-type GEFs and Ect2, and demonstrate that this property enables Ect2 to suppress pulsatile Rho activity dynamics, thereby promoting a stable, non-pulsatile contractile signaling regime during mitosis.